实验小鼠芽囊原虫感染及其对血液指标的影响

doi:10.12140/j.issn.1000-7423.2022.02.019

摘要/Abstract

摘要：

分析实验小鼠芽囊原虫感染及其对小鼠血液指标的影响。2020—2021年,采集石家庄市6个不同实验室饲养的清洁级BALB/c小鼠粪样,提取粪样DNA,PCR扩增芽囊原虫18S特异性片段,测序后在GenBank数据库中进行BLAST比对,分析实验小鼠感染芽囊原虫的亚型,用最大似然法构建基于18S序列的系统进化树。取同批次同环境芽囊原虫检测阳性和阴性小鼠各5只,采血样进行血常规检测,并对检测指标结果进行统计学分析,组间比较采用t检验。共采集小鼠粪样238份,47份PCR扩增出约250 bp的条带,实验小鼠芽囊原虫感染率为19.8%（47/238）。47份阳性样品的序列与GenBank登陆号为MK782501.1（ST1）、MN338076.1（ST2）和MK861934.1（ST7）的序列一致性分别为80.31%~82.79%、97.84%~99.13%和98.79%;系统进化树分析结果显示,阳性样品的序列分别与ST1、ST2和ST7亚型的序列聚集为一支。血常规检测结果显示,芽囊原虫感染小鼠的平均血红蛋白含量为（21.17 ± 0.45）pg,低于未感染小鼠的（23.90 ± 0.60）pg（t = 6.308,P < 0.01）;平均血红蛋白浓度为（480.00 ± 23.64）g/L,低于未感染小鼠的（544.33 ± 15.70）g/L（t = 3.927, P < 0.05）;其余血常规检测指标差异无统计学意义（P > 0.05）。本研究结果提示,实验小鼠感染的芽囊原虫亚型为ST1、ST2和ST7,芽囊原虫感染会引起小鼠部分血液指标异常。

关键词: 实验小鼠, 芽囊原虫, 亚型, 血常规检测

Abstract:

To analyze the infection of Blastocystis and its effect on blood parameters in laboratory mice. Fecal samples of clean-grade BALB/c mice raised in 6 different laboratories in Shijiazhuang City were collected from 2020 to 2021. The fecal DNA was extracted, and then the 18S-specific fragments of Blastocystis were amplified, sequenced and BLAST in the GenBank database. The subtypes of Blastocystis were analyzed, and the phylogenetic tree based on the 18S sequence was constructed by the maximum likelihood method. Five mice infected with Blastocystis and five uninfected mice raised in the same conditions were selected to collect blood samples for the routine blood test. The results were analyzed by t-test to compare the differences between the two groups. A total of 238 fecal samples were collected, and 47 were Blastocystis positive with an infection rate of 19.8% (47/238). The sequence of the 47 positive samples displayed 80.31%-82.79%, 97.84%-99.13% and 98.79% identity to MK782501.1 (ST1), MN338076.1 (ST2) and MK861934.1 (ST7), respectively. The phylogenetic tree showed that the sequences of positive samples were clustered into a branch with the sequences of ST1, ST2 and ST7 subtypes, respectively. The results of the blood test showed that the average hemoglobin content in mice infected with Blastocystis was (21.17 ± 0.45) pg, which was lower than that in the uninfected mice (23.90 ± 0.60) pg(t = 6.308, P < 0.01). The average hemoglobin concentration was (480.00 ± 23.64) g/L, which was lower than that in the uninfected mice (544.33 ± 15.70) g/L(t = 3.927, P < 0.05), while there was no significant difference in other indicators (P > 0.05). The results of this study suggest that the subtypes of Blastocystis which the mice infected were ST1, ST2 and ST7. The infection of Blastocystis will result in abnormal blood parameters in the mice.

Key words: Laboratory mouse, Blastocystis sp., Subtype, Blood routine test

中图分类号:

R531.9

乔海霞, 曹梦娟, 黄晶, 马磊. 实验小鼠芽囊原虫感染及其对血液指标的影响[J]. 中国寄生虫学与寄生虫病杂志, 2022, 40(2): 252-255.

QIAO Hai-xia, CAO Meng-juan, HUANG Jing, MA Lei. Blastocystis infection and its effect on blood parameters in laboratory mice[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2022, 40(2): 252-255.

图/表 3

图1

图2

表1

参考文献 29

[1]	Jiménez PA,, Jaimes JE,, Ramírez JD. A summary of Blastocystis subtypes in North and South America[J]. Parasit Vectors, 2019, 12(1): 376.
[2]	Parkar U,, Traub RJ,, Kumar S, et al. Direct characterization of Blastocystis from faeces by PCR and evidence of zoonotic potential[J]. Parasitology, 2007, 134(pt 3): 359-367.
[3]	Parkar U,, Traub RJ,, Vitali S, et al. Molecular characterization of Blastocystis isolates from zoo animals and their animal-keepers[J]. Vet Parasitol, 2010, 169(1/2): 8-17.
[4]	Dogruman-Al F,, Kustimur S,, Yoshikawa H, et al. Blastocystis subtypes in irritable bowel syndrome and inflammatory bowel disease in Ankara Turkey[J]. Mem Inst Oswaldo Cruz, 2009, 104(5): 724-727.
[5]	Boorom KF,, Smith H,, Nimri L, et al. Oh my aching gut: irritable bowel syndrome, Blastocystis, and asymptomatic infection[J]. Parasit Vectors, 2008, 1(1): 40.
[6]	Souppart L,, Sanciu G,, Cian A, et al. Molecular epidemiology of human Blastocystis isolates in France[J]. Parasitol Res, 2009, 105(2): 413-421.
[7]	Maloney JG,, Molokin A,, da Cunha MJR, et al. Blastocystis subtype distribution in domestic and captive wild bird species from Brazil using next generation amplicon sequencing[J]. Parasite Epidemiol Control, 2020, 9: e00138.
[8]	Alfellani MA,, Taner-Mulla D,, Jacob AS, et al. Genetic diversity of Blastocystis in livestock and zoo animals[J]. Protist, 2013, 164(4): 497-509.
[9]	Stensvold CR,, Traub RJ,, von Samson-Himmelstjerna G, et al. Blastocystis: subtyping isolates using pyrosequencing technology[J]. Exp Parasitol, 2007, 116(2): 111-119.
[10]	Ning CQ,, Ai L,, Hu ZH, et al. Progress of researches on Blastocystis infections in humans and animals in China[J]. Chin J Schisto Control, 2021, 33(1): 95-101. (in Chinese)
[10]	(宁超群,, 艾琳,, 胡主花, 等. 我国人群和动物芽囊原虫感染研究进展[J]. 中国血吸虫病防治杂志, 2021, 33(1): 95-101.)
[11]	Hublin JSY,, Maloney JG,, Santin M. Blastocystis in domesticated and wild mammals and birds[J]. Res Vet Sci, 2021, 135: 260-282.
[12]	Roberts T,, Stark D,, Harkness J, et al. Subtype distribution of Blastocystis isolates from a variety of animals from New South Wales, Australia[J]. Vet Parasitol, 2013, 196(1/2): 85-89.
[13]	Stensvold CR,, Alfellani M,, Clark CG. Levels of genetic diversity vary dramatically between Blastocystis subtypes[J]. Infect Genet Evol, 2012, 12(2): 263-273.
[14]	Stensvold CR,, Clark CG. Pre-empting Pandora’s box: Blastocystis subtypes revisited[J]. Trends Parasitol, 2020, 36(3): 229-232.
[15]	Li Q. Molecular epidemiologic investigation of three important enteric protozoa in birds and mammals from Harbin, Heilongjiang Province[D]. Harbin:Northeast Agricultural University, 2017: 1-60. (in Chinese)
[15]	(黎巧. 黑龙江哈尔滨地区部分鸟类、哺乳动物三种肠道原虫分子流行病学研究[D]. 哈尔滨:东北农业大学, 2017: 1-60.)
[16]	Ramírez JD,, Sánchez LV,, Bautista DC, et al. Blastocystis subtypes detected in humans and animals from Colombia[J]. Infect Genet Evol, 2014, 22: 223-228.
[17]	Menounos PG,, Spanakos G,, Tegos N, et al. Direct detection of Blastocystis sp. in human faecal samples and subtype assignment using single strand conformational polymorphism and sequencing[J]. Mol Cell Probes, 2008, 22(1): 24-29.
[18]	Chai YJ,, Deng L,, Liu HF, et al. First subtyping of Blastocystis sp. from pet rodents in southwestern China[J]. Int J Parasitol Parasites Wildl, 2020, 11: 143-148.
[19]	Li XD,, Zou Y,, Pan J, et al. Prevalence and subtypes of Blastocystis sp. infection in zoo animals in three cities in China[J]. Parasitol Res, 2020, 119(2): 465-471.
[20]	Mohammadpour I,, Bozorg-Ghalati F,, Gazzonis AL, et al. First molecular subtyping and phylogeny of Blastocystis sp. isolated from domestic and synanthropic animals (dogs, cats and brown rats) in southern Iran[J]. Parasites Vectors, 2020, 13(1): 365.
[21]	Betts EL,, Gentekaki E,, Tsaousis AD. Exploring micro-eukaryotic diversity in the gut: co-occurrence of Blastocystis subtypes and other protists in zoo animals[J]. Front Microbiol, 2020, 11: 288.
[22]	Katsumata M,, Yoshikawa H,, Tokoro M, et al. Molecular phylogeny of Blastocystis isolates from wild rodents captured in Indonesia and Japan[J]. Parasitol Res, 2018, 117(9): 2841-2846.
[23]	Cian A,, El Safadi D,, Osman M, et al. Molecular epidemiology of Blastocystis sp. in various animal groups from two French zoos and evaluation of potential zoonotic risk[J]. PLoS One, 2017, 12(1): e0169659.
[24]	Noël C,, Peyronnet C,, Gerbod D, et al. Phylogenetic analysis of Blastocystis isolates from different hosts based on the comparison of small-subunit rRNA gene sequences[J]. Mol Biochem Parasitol, 2003, 126(1): 119-123.
[25]	Yoshikawa H,, Nagano I,, Wu Z, et al. Genomic polymorphism among Blastocystis hominis strains and development of subtype-specific diagnostic primers[J]. Mol Cell Probes, 1998, 12(3): 153-159.
[26]	Li J,, Yuan Y,, Jiang Y, et al. Molecular identification and subtyping of Blastocystis sp. in laboratory rats in China[J]. Parasite, 2020, 27: 35.
[27]	Cheng HS,, Guo YL,, Shin JW. Hematological effects of Blastocystis hominis infection in male foreign workers in Taiwan[J]. Parasitol Res, 2003, 90(1): 48-51.
[28]	Ahmad N. Investigation the role of Blastocystis hominis effect on the levels of Il-10, Il-18 and hematological parameters[J]. Biochem Cell Arch, 2019, 19(2): 3887-3892.
[29]	Li J,, Deng T,, Li XH, et al. The hematology analysis of SD rats infected by Blastocytis hominis[J]. J Gannan Med Univ, 2012, 32(3): 323-324, 337. (in Chinese)
[29]	(李娟,, 邓婷,, 李小花, 等. 人芽囊原虫感染SD大鼠的血常规分析[J]. 赣南医学院学报, 2012, 32(3): 323-324, 337.)

检测指标	未感染小鼠	芽囊原虫感染小鼠	t	P
白细胞/× 10⁹/L	7.03 ± 3.31	7.77 ± 4.46	-0.229	> 0.05
淋巴细胞数/× 10⁹/L	4.45 ± 3.00	3.68 ± 4.01	0.268	> 0.05
单核细胞数/× 10⁹/L	1.62 ± 0.37	1.64 ± 1.28	-0.035	> 0.05
粒细胞数/× 10⁹/L	0.96 ± 0.46	2.44 ± 1.83	-1.357	> 0.05
淋巴细胞百分比/%	59.70 ± 11.82	41.90 ± 28.65	0.995	> 0.05
单核细胞百分比/%	26.60 ± 12.02	20.93 ± 9.64	0.637	> 0.05
粒细胞百分比/%	13.70 ± 0.66	37.17 ± 21.41	-1.897	> 0.05
血红蛋白/g·L^-1	164.93 ± 42.00	186.23 ± 27.06	-0.738	> 0.05
平均血红蛋白含量/pg	23.90 ± 0.60	21.17 ± 0.45	6.308	< 0.01
平均血红蛋白浓度/g·L^-1	544.33 ± 15.70	480.00 ± 23.64	3.927	< 0.05
红细胞/× 10¹²/L	6.92 ± 1.93	8.79 ± 1.43	-1.352	> 0.05
平均红细胞体积/fL	44.07 ± 0.83	44.27 ± 1.42	-0.211	> 0.05
红细胞压积/%	30.40 ± 8.26	39.00 ± 7.60	-1.327	> 0.05
红细胞分布宽度SD/fL	23.33 ± 0.58	24.67 ± 1.53	-1.414	> 0.05
红细胞分布宽度CV/%	13.20 ± 0.10	13.87 ± 0.59	-1.943	> 0.05
血小板/× 10⁹/L	515.33 ± 310.43	770.33 ± 191.25	-1.211	> 0.05
血小板压积/%	0.80 ± 0.45	1.20 ± 0.28	-1.31	> 0.05
平均血小板体积/fL	15.70 ± 0.89	15.53 ± 0.25	0.312	> 0.05
血小板分布宽度/fL	33.23 ± 1.25	33.57 ± 0.35	-0.445	> 0.05
大血小板百分比/%	55.70 ± 7.28	54.30 ± 2.69	0.313	> 0.05